Project Summary Bacterial meningitis is the most common serious infection of the central nervous system (CNS) and a major cause of death and disability worldwide, especially in children. Although antibiotic therapy has changed bacterial meningitis from a uniformly fatal disease to an often curable one, the overall outcome remains unfavorable, with mortality of 5 to 10% and permanent neurologic sequelae occurring in 5 to 40% of survivors, depending on patient age and pathogen. Disruption and dysfunction of the blood-brain barrier (BBB) is a hallmark event in the pathophysiology of bacterial meningitis. Little is known, however, about the very first and crucial interaction between a bacterial pathogen with the BBB that initiates this chain of events, and may ultimately determine a poor or favorable neurological outcome in meningitis patients. This proposal seeks to elucidate the molecular mechanisms of BBB response and function during bacterial infection, and why it fails as a neuroprotective barrier during bacterial meningitis. I hypothesize that BBB disruption may be due to the combined effect of bacterial entry and penetration of brain microvascular endothelium, direct cellular injury by bacterial cytotoxins, and/or activation of host inflammatory pathways that compromise barrier function. These hypotheses will be addressed with both in vitro and in vivo models of BBB penetration using Group B streptococcus as a model human pathogen associated with meningitis. Studies will also utilize isogenic bacterial mutants lacking important virulence factors such as regulators, ?invasins?, and cytotoxins; various knockout and transgenic mice and zebrafish; and key molecular tools to modulate host response pathways in the following specific aims: AIM 1: Characterize the host signaling pathways and bacterial factors that induce tight junction disruption and BBB permeability; AIM 2: Elucidate the mechanisms of bacterial entry and intracellular trafficking for BBB traversal; AIM 3: Characterize the role of astrocytes to BBB function and the pathogenesis of bacterial meningitis. These data will build upon and complement our observations to clarify the crucial position of the brain endothelium in innate immune defense against bacterial pathogens. The knowledge gained as a result of this proposal into the mechanisms of bacterial-brain penetration will provide fundamental and novel insights into BBB function as well as inform treatment strategies for bacterial meningitis and other neurodegenerative disorders.